1. Field of the Invention
The present invention relates to a novel antibacterial compound referred to herein generally as "difficidin", and derivative compounds, and to a process for preparing, isolating, and purifying said compounds. As employed herein, the coined term "difficidin" refers to the compound of Formula I below wherein R.sup.1 is hydrogen, and R.sub.a and R.sub.b have the recited meanings, thus affording various salt forms thereof. Correspondingly, the term "oxydifficidin", as used herein, refers to the compound of Formula I below wherein R.sup.1 is hydroxy, and R.sub.a and R.sub.b have the indicated meanings. The novel antibacterial compounds are obtained by microbiological cultivation of Bacillus subtilis, MB 3575, and MB 4488 deposited with the American Type Culture Collection, Rockville, Md. under the designations ATCC 39374 and 39320, respectively.
The novel antibacterial compounds of the present invention, difficidin and derivative compounds, are broad spectrum antibacterials with good potency against aerobic microorganisms and exceptional potency against anaerobic microorganisms. They also exhibit good in vitro activity against microorganisms that have developed resistance to conventional antibacterials. They may be used parenterally in the treatment of gram positive and gram negative bacterial infections.
2. Brief Description of the Prior Art
Proticin is a known antibacterial compound which has been described as a phosphorous-containing, strongly unsaturated amorphous compound with broad activity spectrum, especially against gram negative pathogens. It is said to have been produced by fermentation of a strain identified as a form of Bacillus licheniformis. A more detailed description of its preparation, characteristics, and antibacterial activity may be found in the following references: (1) Prave, P. et al., J. Antibiotics 25 (1): 1-3, 1972; (2) Vertesy, L., J. Antibiot. 25 (1): 4-10, 1972; (3) Nesemann, G., et al., Naturwissenschaften 59 (2): 81-82, 1972; (4) Ger. Offen. 2,035,812 (Vertesy. L. et al.), "Proticin from Bacillus licheniformis", Farbwerke Hoechst A.-G., July 18, 1970; and (5) Brit. 1,350,271, "Proticin Production by Bacillus licheniformis fermentation," Farbwerke Hoechst A.-G., Apr. 18, 1974. (ref. Chem. Abstracts 81: 48531y, p 30b, 1974).
However, despite the repeated reference to the specific antibiotic compound proticin, the above references fail to characterize any single compound, and the spectral data provided therein is consistent with a number of possible compounds. Since Vertesy Reference (2) above describes the presence of a methyl group at only one of the double bonds, whereas the compounds of the present invention have two such methyl groups, it has been concluded that the novel antibacterial compound of the present invention, wherein R.sup.1 is hydroxy (oxydifficidin) is chemically distinct from the so-called proticin, described in the above references. On the other hand, the novel antibacterial compound of the present invention wherein R.sup.1 is hydrogen (difficidin) is readily distinguished from the compound(s) of the references since it has a different molecular weight.
Chemically, difficidin is a 22-membered macrocyclic polyene lactone phosphate ester with asymmetric centers at C4, C5, C15 and C21, as illustrated by Formula I.
As can be seen, there are two chief compounds within Formula I. One compound is that wherein R.sup.1 is hydrogen, which as already mentioned, shall be referred to hereinafter as "difficidin"; and the other compound is that wherein R.sup.1 is hydroxy, which shall be referred to hereinafter as "oxydifficidin".
Biologically, difficidin has been found to be more potent than oxydifficidin against aerobic microorganisms such as Pseudomonas aeruginosa and Staphyloccoccus aureus. In most cases it was found to be four times more effective against S. aureus, Ps. aeruginosa, Salmonella typhimurium, Proteus, and Serratia marcescens than oxydifficidin. Difficidin and oxydifficidin are also active against a wide spectrum of anaerobic bacteria.